The pKa's of the ionizable amino acids are crucial for the function of many proteins as they are key factors that determine their electrostatic potential and its spatial distribution, often controlling and optimizing enzymatic catalysis. Further, during conformational motions pKa's and protonation states particularly of histidines may change. In established force field simulation, however, this effect is typically not included, and protonation states must therefore be either guessed or derived from experiment. There have been a number of approaches to include protonation effects within simulations, mainly based on continuum electrostatics or implicit solvent molecular dynamics [1--3]. However, these methods lack the effect of the hydrogen bonding and the entropy contribution that comes from the solvent. Here we present the implementation and application of a dynamic protonation atomistic simulation method with explicit solvent, which also allows for explicit solvent constant pH MD simulations, previously developed also in our group [4]. This method is used here to calculate the pKa's of the ionzable groups in proteins. In order to validate it, we selected a number of prototypic proteins and calculated titration curves and pKa values from constant pH simulations at a range of different pH values. The results compare favorably with measured values, and explain atomistically the strong deviations of some of the calculated pKa values from the solution ones. 1. Lee, M. S., Salsbury, F. R., Jr., and Brooks, C. L., III (2004), Proteins 56, 738–752. 2. Khandogin J, Brooks C. L., 3rd., Biophys J. 2005 Jul;89(1):141-57. Epub 2005 Apr, 29. 3. Mongan, J., Case, D. A., McCammon, J. A. J., Comput. Chem. 2004, 25, 2038–2048 4. Donnini S, Tegeler F, Groenhof G, Grubmüller H., J. Chem Theory and Comp 7: 1962–1978 (2011).